KICP
Kinase-Inhibitor Profiling Services
Select the most specific and potent drug candidates for clinical testing
Kinexus presently offers one novel strategy to establish the specificity of putative protein kinase inhibitors.
For more information on our Kinase-Inhibitor Profiling Services please contact our Technical Services representatives toll free at 1-866-KINEXUS or by e-mail at info@kinexus.ca
The selection of the
most specific and potent drug candidates is critical for successful
clinical testing.
At least a third of all pharmaceutical R&D is now focused on protein kinases as drug targets.
More than 536 human protein kinases target the phosphorylation of apparently nearly a million phosphorylation sites in the proteome, of which more than 250,000 have been experimentally confirmed by mass spectrometry.
In view of this, it is critical to establish the specificity of any kinase drug candidate for clinical studies.
On the one hand, the more specific the kinase inhibitors, the lower the chances of off targets that could compromise on the utility of the drug from toxicity and other undesired side-effects.
On the other hand, compounds that more potently inhibit an off target protein kinase could have useful therapeutic applications in diseases that are distinct from those that were originally intended.
Kinexus presently offers only one strategy to establish the specificity of putative protein kinase inhibitors. In the past, we offered radiometric phosphotransferase assays that monitored the ability of 360 different human protein kinases to phosphorylate a protein or peptide with [gamma-32P]ATP or [gamma-33P]ATP.
At least a third of all pharmaceutical R&D is now focused on protein kinases as drug targets.
More than 536 human protein kinases target the phosphorylation of apparently nearly a million phosphorylation sites in the proteome, of which more than 250,000 have been experimentally confirmed by mass spectrometry.
In view of this, it is critical to establish the specificity of any kinase drug candidate for clinical studies.
On the one hand, the more specific the kinase inhibitors, the lower the chances of off targets that could compromise on the utility of the drug from toxicity and other undesired side-effects.
On the other hand, compounds that more potently inhibit an off target protein kinase could have useful therapeutic applications in diseases that are distinct from those that were originally intended.
Kinexus presently offers only one strategy to establish the specificity of putative protein kinase inhibitors. In the past, we offered radiometric phosphotransferase assays that monitored the ability of 360 different human protein kinases to phosphorylate a protein or peptide with [gamma-32P]ATP or [gamma-33P]ATP.
However, with the discontinued availability of Whatman Phosphocellulose paper (which was ideal for such kinase assays) and the increased costs of purchase of these isotopes and radioactivity licensing, it became necessary to adopt other strategies. We tried using the ADP-Glo method from CalBiochem for many years. However, this method has its limitations, which makes it very problematic for screening for protein kinase inhibitors.
Firstly, the ADP-Glo method monitors the production of the ADP product of the kinase reaction rather than the direct phosphorylation of a substrate. Since many kinases also autophosphorylate during kinase assays with added ATP, it is unclear if the production of ADP is from phosphorylation of the substrate or the tested kinase.
Secondly, the ADP-Glo method uses ATP, which is often contaminated with ADP, which means that the signal to noise ratio over the background is often poor, especially if the tested kinase is not very active.
Thirdly, the ADP-Glo method requires three additional reaction steps in which, the remaining ATP is broken down, then the ADP that was produced during the kinase assay is converted back to ATP, and then the ATP is used to drive a luciferase reaction to produce light that is quantified. Since all of these reactions use ATP or ADP as a substrate or product, this could be an issue as many protein kinases inhibitors act competitively with these nucleotides.
Firstly, the ADP-Glo method monitors the production of the ADP product of the kinase reaction rather than the direct phosphorylation of a substrate. Since many kinases also autophosphorylate during kinase assays with added ATP, it is unclear if the production of ADP is from phosphorylation of the substrate or the tested kinase.
Secondly, the ADP-Glo method uses ATP, which is often contaminated with ADP, which means that the signal to noise ratio over the background is often poor, especially if the tested kinase is not very active.
Thirdly, the ADP-Glo method requires three additional reaction steps in which, the remaining ATP is broken down, then the ADP that was produced during the kinase assay is converted back to ATP, and then the ATP is used to drive a luciferase reaction to produce light that is quantified. Since all of these reactions use ATP or ADP as a substrate or product, this could be an issue as many protein kinases inhibitors act competitively with these nucleotides.
In view of these problems, we no longer offer kinase inhibitor or substrate screening services that rely on the ADP-Glo method, and we are working on developing phosphosite-antibody based detection of optimized peptide substrates of kinases. Presently, we are willing to work with clients to develop specific protein kinase assays of interest. However, using our KAM-2025 Antibody Microarray, we have developed another novel method for relatively low cost screening of human protein kinases that uses the endogenous, physiological kinases that are present in lysates from cells.
The Kinexus Kinase Inhibitor Displacement Microarray-based Screen (KIDMAS) is based on the ability of most typical eukaryotic protein kinases to covalently bind to 5-(4-Fluorosulphonylbenzoyl)adenosine (FSBA) at a highly conserved, critical lysine residue located in these enzymes. The binding of a small molecule inhibitor to the active site of these kinases can sterically block the ability of a biotinylated FSBA probe to bind to native protein kinases that are pre-captured on the KAM-2025 Antibody Microarray, which can bind at least 364 different human protein kinases.
Kinexus performs the KIDMAS Service under strict confidentiality, and all materials, information and results are used as directed by the client.
Turnaround with our KIDMAS Service is within 3 weeks of receipt of compounds for testing.
The Kinexus Kinase Inhibitor Displacement Microarray-based Screen (KIDMAS) is based on the ability of most typical eukaryotic protein kinases to covalently bind to 5-(4-Fluorosulphonylbenzoyl)adenosine (FSBA) at a highly conserved, critical lysine residue located in these enzymes. The binding of a small molecule inhibitor to the active site of these kinases can sterically block the ability of a biotinylated FSBA probe to bind to native protein kinases that are pre-captured on the KAM-2025 Antibody Microarray, which can bind at least 364 different human protein kinases.
Kinexus performs the KIDMAS Service under strict confidentiality, and all materials, information and results are used as directed by the client.
Turnaround with our KIDMAS Service is within 3 weeks of receipt of compounds for testing.